The Australian Rules of Harness Racing states
that a horse shall be presented for a race free of prohibited substances. Many
registered, unregistered and illegal medications, substances and their
metabolites are listed as prohibited substances under these Rules. A smaller
number of naturally occurring substances is listed as being prohibited if a
threshold level of the substance in a horse’s system is exceeded. Plasma Total
Carbon dioxide (TCO2) concentration is one such substance.

The TCO2 concentration is the total amount of
carbon dioxide that can be liberated from blood plasma. Carbon dioxide occurs
naturally in the blood of horses at concentrations that vary between horses. A
number of physiological factors can affect a horse’s TCO2. These factors
include the horse’s sex and age, diet, water intake, time of day, climate and
ambient temperature and exercise. Pathological conditions such as respiratory
disease can also affect TCO2.

Alkalising Substances

Alkalising substances, including bicarbonate
and related substances, can increase plasma TCO2. Exercise physiologists advise
that the administration of these substances may assist the horse recovering from
strenuous exercise. A large number of proprietary oral preparations containing
alkalising substances are promoted to the horse industry. Alkalising substances
can be used as buffering or bulking agents in other horse treatments.

TCO2 threshold

In the early 1990’s, the practice of ‘milkshaking’
came to the notice of racing authorities. The milkshake consisted of sodium
bicarbonate, glucose and a number of other substances and was given to the horse
via a stomach tube. Trainers believed that this practice improved the athletic
endurance of the horse. In some cases, the ‘milkshake’ could cause colic,
dehydration and diarrhoea and therefore be detrimental to the horse’s
performance.

There was some concern that ‘milkshaking’
could change the pH of the urine and either inhibit or increase the excretion of
certain drugs, reducing the chance of detection in a post race urine sample.

In an attempt to control this practice, a
TCO2 threshold was set by the Australian Harness Racing Council. Horses with a
TCO2 concentration above this threshold were considered to have received an
excess of alkalising agent, whether as a milkshake or by another method, for the
purpose of affecting the performance of the horse.

Initially the TCO2 threshold was set at
37mmol/l. After reviewing the actual TCO2 concentrations in racing standardbreds
over a three-year period, the threshold was subsequently lowered to 35mmol/l in
1995.

Because of the laboratory method employed for
measuring TCO2 concentration, each test result is reported as a range rather
than a single result. The limits of the range are determined by the laboratory
uncertainty of measurement. All results are reported as TCO2 result +/-
uncertainty of measurement. A single uncertainty value is applied to all
measurements even though the uncertainty of measurement can increase with the
measurement of increasing TCO2 concentrations. The Australian Racing
Laboratories set the uncertainty measurement at 1.2mmol/l to 1.4mmol/l.

In the last 18 months, there was concern
raised by trainers and their advisers that an increasing number of horses were
exceeding the 35mmol/l threshold. The penalty imposed on most violations of the
Rule was a 12-month disqualification.

The majority of the violations appeared to be
occurring in one Australian State (South Australia). There was considerable
debate about the reasons for the TCO2 concentration increase including
physiological effects, such as pre race excitement elevating TCO2
concentrations, and doubts about the laboratory testing process for TCO2.

TCO2 Review

The Australian Harness Racing Council created
a subcommittee to review the TCO2 situation in Australia under the following
Terms of Reference:

Level of laboratory uncertainty factor in the testing
process

Integrity of the process

The mean TCO2 level of the harness racing population

The mean level of untreated harness racing population

Has the mean level increased and why?

There were three information-gathering
strategies available to the subcommittee.

Submissions from technical experts and the general
harness racing community. 30 technical and general submissions were
received. A number of published and unpublished papers were submitted.

Biometrical analysis by an independent biometrical
service of all pre race TCO2 data since 1995. Approximately 60,000
individual pre race TCO2 results from all Australian states were analysed.

A mail survey of the feeding practices of the
Australian Standardbred. A total of 525 trainers returned information on the
feeds and additives given to the horses in their stable.

The subcommittee’s findings on TCO2 can be
categorised under three main headings:

Sampling influence

Laboratory influence

Trainer influence.

Sampling influence

The collection and storage of the blood
sample for TCO2 testing is critical for the accuracy of the test result. Any
variation in the collection method can affect the TCO2 concentration. In most
cases, adverse handling of the blood sample will decrease the TCO2
concentration. These handling variations that can occur include:

the period of time between blood collection and race
start,

the number of blood samples collected from each horse,

the anti-coagulant used,

the size of the blood tube,

the fill of the tube during collection,

the handling and storage of the tubes after collection

the transport to the laboratory

the period of time between blood collection and receipt
and testing at the laboratory.

All Australian States now effectively have
the same protocol for collection of blood samples at the racetrack. The horses
are sampled before they are exercised or ‘warmed-up’ and within 90 minutes
before race start.

A Subcommittee has been formed to further
review and refine the sampling protocol.

Laboratory influence

The type of equipment used to measure the
TCO2 concentration at the laboratory should be consistent between laboratories.
At present, all Australian Racing Laboratories use the Beckman Synchron ELISE to
measure TCO2 concentration.

The protocol for analysing the blood sample
by the equipment should be the similar between laboratories. All Australian
laboratories use the Australian Scientific Enterprise Pty Ltd (ASE) standards to
calibrate the Beckman to the range of TCO2 concentrations that can occur in the
racehorse. A control sample (Verichem) is used to determine the effectiveness of
the machine calibration before and during each period of sample testing.

The laboratories are currently reviewing the
uncertainty of measurement and it may change in the future.

The Subcommittee is collaborating with the
laboratories to further refine the current protocol and make recommendations on
any future improvements.

Trainer Influence.

The Diet Survey revealed that ongoing
education of the trainer and their advisers on the effective use of feed
additives was essential.

The majority of trainers used a variety of
products containing alkalising agents yet did not recognise that these products
contained substances that could increase the TCO2 concentration of their horses.
The multiple use of registered and unregistered products containing these
substances in the horse was evident. The effect of these practices on the horse’s
resting TCO2 is not known.

The practice of withdrawing water from horses
for a period of time before race start was reasonably common. This practice can
increase TCO2 concentration. The combined effect of adding alkalising substances
to feed and pre-race withdrawal of water could increase the horse’s TCO2
towards the threshold limit.

An equine nutritionist is reviewing the
results of the Diet Survey and is developing recommendations that could be
incorporated into an education program on the knowledge of feeding practice sand
understanding of feed additives for trainers and their advisers.